Diminished ability of thymus to produce naive T cells with progressive aging remains a fundamental and puzzling phenomenon for immunology and to-date, an intractable clinical condition that contributes to immune dysfunction in elderly. With advancing age, the thymus undergoes striking fibrotic and fatty changes that culminate in its transformation into adipose tissue. The lineage of ectopic adipocytes and the mechanism of development of thymic adipocytes during aging are not well understood. Using lineage-tracing, our research team has shown that FoxN1+ thymic epithelial cells (TECs), that are necessary for T cell development, can transition and give rise to thymic adipogenic precursors. Consistent with this new paradigm, our latest preliminary data provide further in vivo evidence that proadipogenic cells in aging thymus can originate from epithelial and endothelial lineages via a secondary mesenchymal precursor. Based on our novel findings, the central hypothesis of this proposal is that the transition of TECs and endothelial cells contribute towards the lineage of thymic adipocytes and leads to age-related thymic involution. The corollary is that blocking the fibrogenesis and pro-adipogenic signaling in TEC and endothelial lineage cells will protect against aging of thymus. Experiments in Aim 1 will test the hypothesis 1 that with advancing age transition of FoxN1+ cells into adipogenic precursors via epithelial-mesenchymal transition (EMT) process gives rise to ectopic thymic adipocytes and compromises the thymic stromal microenvironment. Aim 2 will test the hypothesis 2 that that endothelial- lineage cells serve as adipogenic progenitors and will reveal that blocking the ectopic adipocyte development in thymic perivascular space protect against thymic aging. The Aim 3 will test the hypothesis 3 that age-related fibrogenesis in thymus causes reduction in thymopoiesis and reveal that mechanisms that commit secondary mesenchymal cells into adipocytes participate in thymic dysfunction. The long-term goal of this research project is to understand the mechanisms that cause thymic adiposity and to develop new approaches to prevent or reverse the process of age- related thymic involution.